Angiogenesis, the outgrowth of new capillaries from pre-existing vessels, is essential for embryonic development, organ formation, tissue regeneration, and remodeling [Folkman, J. & Shing, Y. (1992) J. Biol. Chem. 267, 10931-10934]. It also contributes to the development and progression of a variety of pathological conditions, including tumor growth and metastasis, cardiovascular diseases, diabetic retinopathy, rheumatoid arthritis, and psoriasis [Folkman, J. (1995) Nat. Med. 1, 27-312]. Angiogenesis and vasculogenesis are complex multistep processes that include proliferation, migration and differentiation of endothelial cells, degradation of the extracellular matrix, tube formation, and sprouting of new capillary branches [Hanahan, D. & Folkman, J. (1996) Cell 86, 353-364; Risau, W. (1997) Nature (London) 386, 671-674]. The complexity of the angiogenic processes suggests the existence of multiple controls of the system, which can be transiently switched on and off. A switch of the angiogenic phenotype in tissues is thought to depend on a local change of the balance between angiogenic stimulators and inhibitors [Folkman, J. (1995) N. Engl. J. Med. 333, 1757-1763].
Among many described angiogenic factors, vascular endothelial growth factor (VEGF)/vascular permeability factor is one of the best-characterized positive regulators with its distinct specificity for vascular endothelial cells [Senger, D. R., Galli, S. J., Dvorak, A. M., Perruzzi, C. A., Harvey, V. S. & Dvorak. H. F. (1983) Science 219, 983-985; Ferrara, N. & Henzel, W. J. (1989) Biochem. Biophys. Res. Commun. 161, 851-858; Gospodarowicz, D., Abraham, J. A. & Schilling, J. (1989) Proc. Natl. Acad. Sci. USA 86, 7311-7315]. The biological actions of VEGF include stimulation of endothelial cell proliferation, migration, differentiation, tube formation, increase of vascular permeability, and maintenance of vascular integrity [Mustonen, T. & Alitalo, K. (1995) J. Cell Biol. 129, 895-898; Ferrara, N. & Davis-Smyth, T. (1997) Endocr. Rev. 18, 4-25; Thomas, K. (1996) J. Biol. Chem. 271, 603-606; Risau, W. (1997) Nature (London) 386, 671-674; Breier, G. & Risau, W. (1997) Trends Cell Biol. 6, 454-456]. The angiogenic responses induced by VEGF are mediated by tyrosine kinase receptors, which are expressed primarily on vascular cells of the endothelial lineage [Mustonen, T. & Alitalo, K. (1995) J. Cell Biol. 129, 895-898; De Vries, C., Escobedo, J. A., Ueno, H., Huck, K., Ferrara, N. & Williams, L. T. (1992) Science 255, 989-99; Terman, B. I., Dougher-Vermazen, M., Carrion, M. E., Dimitrov, D., Armellino, D. C., Gospodorawicz, D. & Bohlen, P. (1992) Biochem. Biophys. Res. Commun. 187, 1579-1586].
Inhibition of cell adhesion to the endothelial cell membrane (ECM), the fundamental step for activation, survival, targeting and migration of activated endothelial cells, might be one of the most promising target mechanisms for anti-angiogenesis. Not only VEGF is involved in these mechanisms but many of these interactions are also mediated by integrins, a family of multifunction cell adhesion receptors. Members of the integrin family are non-covalently alpha/beta heterodimers that mediate cell-cell, cell-extracellular matrix and cell-pathogen interactions. Until now, 19 different integrin alpha subunits and 8 different beta subunits are known that combine to form at least 25 different alpha/beta heterodimers with different ligand specificity. The ligands for the extracellular domain of many integrins are the proteins of the extracellular matrix and the intracellular domain of the integrins are either directly or indirectly connected to intracellular components such as kinases and the cytoskeleton. Integrins serve as bidirectional signalling receptors, whereby protein activities and gene expression are changed by integrins in response to ligand binding to the extracellular domain thereof, which is also referred to as outside-in-signalling. On the other hand, the affinity of the integrins is modulated in response to intracellular changes such as binding of proteins to the extracellular domain of the integrin, which is referred to as inside-out signalling [Humphries (2000) Biochem Soc Trans. 28, 311; Hynes (2002) Cell, 110, 673].
Several studies on the integrin pattern on activated endothelial cells, mice gene knockouts and inhibition studies in angiogenic animal models with antibodies, peptides and small molecules have provided information about integrins and ECM proteins involved in critical steps of angiogenesis [Brooks (1994) Science, 264, 569; Brooks (1996) Eur J Cancer, 32A, 2423; Mousa (2002), Curr Opin Chem Biol, 6, 534; Hynes (2002) Nature Medicine, 8, 918; Kim (2000) Am J Pathol, 156, 1345]. From this work it appeared that the fibronectin receptors alpha-v-beta-3, alpha-v-beta-5 and the fibronectin receptor alpha5beta1 play a critical role in angiogenesis. Alpha5beta1 expression is significantly upregulated in blood vessels in human tumors and after stimulation with growth factors and, once expressed, alpha-5-beta-1 regulates the survival and migration of endothelial cells in vitro and in vivo.